A brief interaction with a stranger is an important first step in the formation of a new social relationship. During this brief interaction, two individuals typically evaluate one another to decide whether or not to move forward and form a relationship. Social evaluation may be perceived as mildly threatening and therefore be psychologically stressful. Differences in personality traits may moderate how individuals respond to psychosocial stress and how stress affects the subsequent social interaction between them. This study investigated the role of stress and personality in the early stages of social relationship formation in the laboratory using the Trier Social Stress Test (TSST) as experimental paradigm to induce psychosocial stress from social evaluation threat. Participants were 156 heterosexual men and women randomly assigned to either a TSST or a control condition. All participants later engaged in a brief social interaction with an opposite sex confederate. In addition, changes in self-reported personality before and after all manipulations were assessed to investigate whether stress and social interaction changes how an individual perceived oneself. There was no significant moderating effect of personality on cortisol stress responses, but a significant moderating effect of gender. Specifically, males displayed greater changes in cortisol levels than females, while females reported greater changes in anxiety scores than males after TSST. In male participants, higher Openness to Experience was associated with lower testosterone responses to social interaction regardless of treatment. All participants except TSST females reported higher extraversion score, only control females reported higher emotional stability score, and only TSST females reported lower conscientiousness score.